Swainsonine, 1, (the general structure of which is presented in FIG. 1) is an alkaloid of molecular weight 173.1 which may be derived from both natural and synthetic sources. Swainsonine is a competitive inhibitor of Golgi alpha-mannosidase II (Molyneux, R. J. et al., Science (Wash. D. C.) 216:190 (1981); Tulsiani, D. R. P. et al., J. Biol. Chem. 257:7936 (1982)), an enzyme important in the oligosaccharide processing pathway leading to the synthesis of complex membrane oligosaccharides and secreted glycoproteins. This action, as well as the immunomodulatory effects of swainsonine, accounts in part for antimetastatic and antineoplastic activity in murine tumor model systems. Swainsonine has also been shown to mediate a diverse array of effects, which include inhibition of tumor growth (Dennis et al., J. Nat'l. Cancer Inst., 81:1028 (1989)) and metastasis (White et al., Anticancer Res., 10:1515 (1990)), synergistic with poly-IC and interleukin-2 (J. W. Dennis, Mt. Sinai Hospital, European Patent Application 87308605.2); augmentation of natural killer (Inamura et al., J. Antibiot. Tokyo, 38:936 (1985)) and macrophage (Newton et al., Cancer Commun., 1:373 (1989)) tumoricidal activity; induction of cytokine synthesis and secretion (Newton et al., Cancer Commun., 1:373 (1989)); enhancement in expression of lak (Newton et al., Cancer Commun., 1:373 (1989)) and HLA class 1 specific antigens (White et al., Anticancer Res., 10:1515 (1990)); activation of protein kinase C (Breton et al., Cancer Commun., 2:333 (1990)); stimulation (5-10 fold increase) of bone marrow (BM) proliferation (White et al., Cancer Commun., 3:83 (1991)); engraftment efficiency, and colony-forming unit activity (CFU-GM, CFU-GEMM, and BFU-E) as assessed by both in vivo and in vitro assays. Swainsonine has also been shown to confer protection against toxicity induced by high dose chemotherapy, to stimulate BM cell proliferation, and to accelerate recovery of BM cellularity when used in combination with chemical agents commonly used in treatment of human malignancies, (J. Nat. Cancer Inst., 83:1125, 1991). Published European Patent Application 0 104 826 also discloses swainsonine; moreover, it indicates therapeutic dosages and how they are determined for use in treating diseases accompanied by depressed immunoactivity.
Swainsonine offers a number of advantages over recombinant growth factors and cytokines. It is possible that swainsonine induces the simultaneous paracrine production, in modest quantities, of a number of cytokines. This effect may allow marrow protection in the absence of toxicity and the complexities involved in administration of one or more growth factors to an individual. In addition, swainsonine has biological activity when administered orally. Therefore, the potential exists for a convenient route of administration of swainsonine which is not possible using recombinant proteins. Furthermore, the simplicity of the molecule may facilitate inexpensive production of a suitable formulation.
With regard to the syntheses of (-)-swainsonine, several enantioselective syntheses of the natural alkaloid have been reported. Many of these utilize carbohydrates as chiral precursors. Syntheses from D-mannose [Gonzales et. al., Bull. Chem. Soc. Jpn., 65: 567 (1992); T. Takaya et al., Chem. Lett., 1201 (1984); G. W. J. Fleet et al., Tet. Lett., 25: 1853 (1984); G. W. J. Fleet et al., Tetrahedron, 43: 3083 (1987); G. W. J. Fleet et al., Tet. Lett. 30: 7261 (1989)], D-glucose [A. C. Richardson et al., Carb. Res. 136: 225 (1985); A. C. Richardson et al., J. Chem. Soc., Chem. Comm. 447 (1984); T. Saumi et al., Chem. Lett. 513 (1 984); T. Saumi et al., Carb. Res., 136: 67 (1985))], D-erythrose [J. K. Cha et al., J. Amer. Chem. Soc., 111: 2580 (1989); W. H. Pearson, Tet. Lett., 52: 7571 (1990)], D-xlyxose [A. R. Chamberlain, J. Amer. Chem. Soc., 112: 8100 (1990)], D-ribonolactone [N. Ikota et al., Chem. Pharm. Bull., 36: 1143 (1988)], glutamic acid [N. Ikota et al., Chem. Pharm. Bull., 35: 2140 (1987); N. Ikota et al., Chem. Pharm. Bull. 38.2712 (1990); D-tartaric acid [D. J. Hart et al., J. Org. Chem., 53: 6023 (1988)] have all been reported. The synthesis of (-)-swainsonine has also been reported starting from achiral precursors. K. B. Sharpless [K. B. Sharpless et al., J. Org. Chem., 50: 422 (1985)] reported a 21 step synthesis starting from trans-1,4-dichloro-2-butene employing the methodology of the Masamune/Sharpless iterative approach to polyhydroxylated natural products [S. Masamune, K. B. Sharpless et al., J. Org. Chem., 47: 1373 (1982), Science, 220: 949 (1983)].
Castanospermine, 2,1S-8a-beta-octahydroindolizine-1-alpha-6-beta-7-alpha-8-beta-tetraol (see Formula 2 below) an inhibitor of the endoplasmic reticulum enzyme alphaglucosidase, has been converted specifically to mono- and di-O-acylated derivatives (P. S. Liu et al., Tet. Lett, 32:719 (1991); P. S. Liu et al., Tet. Lett., 31:2829 (1990 ); A. L. Margolin et al., J. Amer. Chem. Soc., 112:2850 (1990): W. K. Anderson et al., Tet. Lett., 31:169 (1990)). It has been reported that several O-acyl derivatives of castanospermine are as much as 20 times more active than castanospermine itself in inhibiting HIV replication (P. S. Sunkara et al., The Lancet, 1206 (1989)). The structure of castanospermine is illustrated in FIG. 2.
The lipophilicity of a hydrophilic drug can enhance absorption from the GI tract and alter the organ distribution. Rall et al., (in Ann. Rev. Pharm., 2:109 (1962)), have reported that small lipophilic molecules cross the blood-brain barrier more efficiently than large ones. Esters of hydrophilic drugs may be cleaved by enzymes in the blood stream releasing the free drug, thus the profile of excretion may also be affected by the structure of these derivatives. An example of this is acetylsalicylic acid, which is cleaved to the active agent, salicylic acid, in the liver and in various other tissues (D. Lednicer et al., The Organic Chemistry of Drug Synthesis, J. Wiley and Sons, Vol. 1, pg. 108, 1977).
A number of acyl derivatives of swainsonine have been synthesized. In the course of the initial structure determination of swainsonine, acetylation of swainsonine with acetic anhydride at room temperature was found to yield a 1, 2-diacetate derivative (S. M. Colegate et al., Aust. J. Chem., 32:2257 (1979)). Researchers at Fujisawa Pharmaceutical Co., Ltd. have also prepared a number of acylated derivatives of swainsonine as immunomodulators (Jpn. Kokai Tokkyo Koho 61,277685). These compounds include tri-O-acylated derivatives of swainsonine, 1, 2-di-O-acetylated derivatives of swainsonine, 1, 8-di-O-benzoyl swainsonine and 2, 8-di-O-benzoyl swainsonine and 8-O-acylated derivatives. Other acetylated derivatives of swainsonine reported in the chemical literature arise as intermediates in various syntheses of swainsonine. These are tri-O-acylated compounds.
Because swainsonine contains three similar hydroxyl functions (See FIG. 1), direct modification of swainsonine by manipulation of these hydroxyl groups represents a formidable synthetic challenge. Japanese Kokai Tokyo Koho 61,277685 discloses the difficulty in differentiating between the 1 and 2 hydroxyl groups of swainsonine.
The reaction of 1, 2-diols with dibutyltin oxide results in the formation of a cyclic 5-membered dibutylstannyl derivative. These have been successfully used in carbohydrate and nucleoside chemistry (D. Wagner et al., J. Org. Chem., 39:24 (1974)) as a protecting group or as an activating group for subsequent alkylations, acylations and oxidations (Chem. Pharm. Bull., 37:2344 (1989)).
Other than the initially reported conversion of swainsonine to its 1, 2-di-O-acetyl derivative, site specific acylation of swainsonine is not known to have been reported to date.
The reaction of diols with dibutyltin oxide is known to form a cyclic dibutylstannane derivative which can function as both a protecting group, or be utilized to activate one of the hydroxyl groups toward alkylation, acylation (J. Org. Chem. 39: 24 (1974)), or even oxidation (Chem. Pharm. Bull, 2344 (1989). This technique has been utilized in carbohydrate, and nucleotide chemistry where the diol is part of a molecule containing other hydroxyl groups. It was also utilized to prepare esters of castanospermine (Tetrahedron Letters, 169 (1990).
The glycosylation of tin alkoxides was reported in 1976 [Carbohydrate Res. 51: C13 (1976)]. The reaction of acetobromoglucose with the tributyltin salts of various alcohols, in 1,2-dichloroethane was catalyzed with tin tetrachloride. These reactions afforded reasonable yields of the glycosides. When the reaction is run with tetraethylammonium bromide, orthoesters are produced. The conversion of such orthoesters into 1,2-trans-glycosides is well established [Kochetkov et. al., Tetrahedron 23: 693 (1967); Zurabyan et. al., Carbohydrate Res. 26: 117 (1973); T. Ogawa et. al., Tetrahedron 37: 2787 (1981); T. Ogawa et. al., Tetrahedron 37: 2787 (1981)]. These papers deal with the glycosylation of the salts produced by reaction of trialkyltin enolates with glycosyl halides.
Glycosyl donors are commonly carbohydrates activated for coupling to hydroxyl groups. This includes activation as the glycosyl bromide, chloride, fluoride, tosyl, or oxazoline compound or activation via imidate chemistry [B. Wegmann et. al., J. Carb. Chem., 357: (1987); R. R. Schmidt, Tet. Lett. 32: 3353 (1991)] or as a thioglycoside [J. O. Kihlberg, et. al. J. Org. Chem. 55, 2860 (1990)], and cited references)]. The reaction of glycosyl donors with hydroxyl groups to afford glycosides is normally promoted utilizing silver salts, mercury salts, trifluoromethanesulfonic anhydride [H. P. Wessel, Tet. Lett. 31: 6863 (1990)].